The virus denominated human Adenovirus (ADV) is responsible of producing diverse clinical pictures in the population, which include from conjunctivitis, gastroenteritis to respiratory diseases of all types (such as mild pharyngitis to acute respiratory diseases of the lower respiratory tract, such as pneumonia). Studies have demonstrated that this virus produces a high percentage of morbidity and even the death of infants, military recruits and immunocompromised people worldwide. ADV is known for being a persistent virus remaining latent in previously infected individuals and it can be reactivated by diverse factors, carrying out to the generation a more severe disease. Its genome contains double stranded DNA, which does not have nuclear envelope and codifies for 9 proteins. The capsid that covers it has icosahedron form and it is composed by 3 main proteins: Hexon, Penton (highly conserved) and the fiber. Studies have reported that the penton protein and the fiber are in charge of the attack and entry to the host cell, promoting this last one the internalization of the virus interacting with the receptors in the cell surface. There are more than, 50 serotypes of ADV in humans, which are grouped in 6 species which are subdivided from A-F. Within the subdivision some of the strains—B, C and E—have greater clinic relevance in respiratory diseases, wherein the severity grade of the disease will depend on the age (children <4 years and adults) and the immunologic status of infected patients.
Nowadays the diagnostic and detection of ADV methods performed by the Public Health services include a test based on the viral antigen detection by direct immunofluorescence in swab or nasopharyngeal aspirate samples, polymerase chain reaction (PCR) and cell culture, from a blood sample. Of the mentioned techniques, the viral panel based on immunofluorescence allows to detect a greater number of respiratory virus, being 12 types in the case of Respiratory Viral Panel (RVP) PCR utilizing Luminex xTAG and 14 types for the eSensor Respiratory Viral Panel. Despite to this wide range of detection, is important to note the cost and response time factors used by them (in the case of the first one the cost is approximately USD 60-80 (CLP 42.000-56.000) and a response time between 12 to 18 hours).
Based on the above background, it is fundamental generating and counting with a diagnostic test ex vivo or fast in vitro, effective and low cost for the ADV detection which can compete against the features of the available diagnostic methods. In front of such problem, the monoclonal antibodies of the invention appear as an alternative needed to address said need, since they allow the specific recognition of viral antigens in samples of patients infected with ADV. Currently, there are antibodies used for research (Thermo Scientific Cat. #MS-587) and diagnostic purposes (Abcam® ab6982), however, monoclonal antibodies which specifically recognize the pIII protein of this virus have not been reported.
A monoclonal antibody is a type of homogeneous antibody which is characterized by specifically recognizing a single antigen. They are produced by a single hybrid cell (hybridoma), which is product of the fusion of a B lymphocyte clone and a tumor plasmatic cell. The property of specifically binding and with high affinity to an antigen has driven the development of monoclonal antibodies as a tool of great utility for the detection of molecules which generate a great scientific, clinical interest and of industrial use. Nowadays, the monoclonal antibodies are widely used, in both basic and applied research, due to their specificity and reproducibility, which allow to support the research in better manner. However, it is in the biomedicine area wherein the monoclonal antibodies have had huge practical applications, either for diagnostic and treatment of multiple infectious diseases, and as therapy for other pathologies. While it is true that the monoclonal antibodies are used in all types of detection and diagnostic techniques, it is in the design of kits for in vitro diagnostic wherein the best results have been obtained. For that, there are currently diverse fast detection kits, such as the pregnancy test, which is based on the determination of the chorionic gonadotropin (hCG) levels in the urine using anti hCG antibody. Also, the monoclonal antibodies for therapeutic use have gained great relevance. Currently, there are therapeutic treatments for different pathologies, using commercial monoclonal antibodies such as: Alemtuzumad, Gemtuzumab ozogamicina, Rituximab, Trastumab, among others.
Currently, there are monoclonal antibodies against different proteins of the adenovirus: however, it is highly noted the production of antibodies against the hexon protein and the capside fiber in several published works. A clear example of the above, it is observed in U.S. Pat. No. 4,487,829, wherein they achieve to generate antibodies called anti 2-Hx-2 capable of detecting the hexon antigen in the C subgroup (1,2,5) of human adenovirus. On the other hand, the present application shows the existence of new monoclonal antibodies, or fragments thereof, which recognize the pIII protein of the Adenovirus (ADV).
Additionally, document US20110262892A1 describes detection or virus measurement methods, wherein one of the detected virus is Adenovirus, however, this document does not describe specific monoclonal antibodies or fragments thereof which recognize the Adenovirus pIII protein as the present application does.
Having into account the high production and competitiveness in the market of anti hexon monoclonal antibodies, it is required the search and development of new monoclonal antibodies against conserved and abundant proteins in the virion structure. Such is the case of the monoclonal antibodies of this invention, which are directed against the pIII protein (base penton), a highly conserved protein among ADV subgroups. These are the first monoclonal antibodies produced against this protein, which demonstrated being capable of detecting viral particles from the clinical samples with high sensitivity.
Thus, the present invention comprises products, such as monoclonal antibodies, and an alternative method which uses them for detection and fast, effective and accurate diagnostic in patients infected with ADV with a 100% of specificity in clinical samples and capable of detecting by ELISA, concentrations equivalent 1.5 ng of the specific antigen. This will allow clinical professionals determining in an early manner an appropriate clinical protocol for individuals suffering a respiratory infection caused by this virus.